The technique of equations 1 and 2 works well when the peaks are well separated, as with Figure 2a, where R s = 1.3. Because the half-height peak width is easier to measure (no tangent drawing involved), most data systems use the half-height method (equation 2) to calculate resolution. The peak width at the baseline for a Gaussian peak is 4σ (4 standard deviations), whereas at the half-height, it is 2.354σ, so the factor in equation 2 is (2 × 2.354/4) = 1.18. That is, the resolution is the difference in retention times divided by the average baseline peak width (thus the factor of 2 in equation 1). Where w b1 and w b2 are the baseline peak widths between tangents drawn to the sides of the peaks, and w h1 and w h2 are the corresponding peak widths measured at half the peak height. Most of us use the method of equation 1 or 2 to calculate the resolution, R s, of a pair of peaks with retention times t 1 and t 2: In this month's "LC Troubleshooting" installment, I would like to share a simple technique to estimate resolution that has been in use for many years (for example, see reference 1), but may not be well known because of our dependence on automatic data processing systems today.įigure 1: An example of poorly resolved peak pairs A, B, and C. In each case, the valley between the peaks does not dip below 50% of the height of the smaller peak, making it impossible to measure the peak width at the baseline or half-height. An example of this is shown for peak pairs A, B, and C in the chromatogram of Figure 1. I have had several reader inquiries lately regarding how to estimate resolution between two peaks in a liquid chromatography (LC) separation when the traditional calculation doesn't work. How can resolution be determined when peak width cannot be measured?
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